E-Poster Presentation ESA-SRB-ANZBMS 2021

Identifying the cell cycle-related factors in the pregranulosa cells driving primordial follicle activation (#541)

Emily R Frost 1 2 3 4 , Gunes Taylor 4 , Stefan Boeing 5 6 , Mark A Baker 2 3 , Robin Lovell-Badge 4 , Jessie M Sutherland 2 3
  1. Sex Development Lab, Hudson Institute for Medical Research, Oakleigh East, VIC, Australia
  2. Priority Research Centre for Reproductive Science, School of Biomedical Sciences and Pharmacy, University of Newcastle, Callaghan, NSW, Australia
  3. Pregnancy and Reproduction Program, Hunter Medical Research Institute, New Lambton Heights, NSW, Australia
  4. Stem Cell Biology and Developmental Genetics Lab, The Francis Crick Institute, London, United Kingdom
  5. Scientific Computing – Digital Development Team, The Francis Crick Institute, London, United Kingdom
  6. Bioinformatics and Biostatistics Facility, The Francis Crick Institute, London, United Kingdom

Female fertility is controlled by the number of oocyte-containing follicles within the ovary. These follicles (termed primordial follicles) are established early in development and must sustain reproductive function across a lifespan. Primordial follicle activation is the first step in the selective development of mature follicles for ovulation. The mechanisms that control the activation of primordial follicles are still largely unknown. We used single-cell RNA sequencing to examine the transcriptional profile of mouse embryonic and neonatal ovaries at three timepoints: embryonic day (E) 18.5, postnatal day (PND) 4 and PND7, which correspond with primordial follicle formation and activation. In total, 24,810 cells were sequenced, and we identified 5 distinct clusters of granulosa cells. We distinguished the transcriptomic signature of pregranulosa cells during follicle activation in granulosa cell cluster 1 (Gc_1), by comparing with the activated granulosa cells present in granulosa cell cluster 4 (Gc_4). 389 genes were upregulated in the Gc_1 cluster, compared with 278 genes in Gc_4, and these differentially expressed genes were annotated to divergent cell cycle processes. To validate the role of cell cycle in regulating follicle activation, we performed transcriptomic analysis of a mouse line deficient in Cdkn1b/p27kip1 (a cell cycle inhibitor), which shows a follicle activation phenotype (Rajareddy et al.2007). The gene expression signature identified in pregranulosa cells in our single cell RNA sequencing analysis was observed precociously in the infertile Cdkn1b-/- mouse line. Combined, this data suggests that a loss of cell-cycle regulation in pregranulosa cells results in precocious primordial follicle activation.

This dataset provides the groundwork for characterising a gene regulatory network that regulates primordial follicle activation. An understanding of the factors that act co-operatively to stimulate/inhibit follicle activation will reveal key insights into how the ovarian reserve is established and how this may be dysregulated in female infertility disorders and premature reproductive decline.